Fig. 9

Spautin-1 prevents the autophagy-associated osteoblastic differentiation of Human Gingival Mesenchymal Stem cells. a HGMSCs were plated on sterile coverslips, let adhere and cultured for 21 days in control medium or in differentiation medium supplemented with 1 μM resveratrol (Diff + RV) in the absence or the presence of spautin-1 (Sp1). The coverslips were then fixed and processed for immunofluorescence staining of the autophagy marker LC3 and of the osteoblastic differentiation marker OCN. As an index of autophagy in the cells, LC3 puncta were quantified (as per the guidelines 36). b HGMSCs were plated and treated as in a and the coverslips processed for immunofluorescence staining of the osteoblastic differentiation markers OCN and COL1A1. Integrated fluorescence intensity quantified with the ImageJ software is shown in the histograms. c HGMSCs cells were plated on plastic and treated as in a and at the end the cultures were processed for Alizarin Red S staining of extracellular calcium deposits. The stained area was quantified using the ImageJ software